Birth of mice after in vitro fertilization using C57BL/6 sperm transported within epididymides at refrigerated temperatures

The transportation of cryopreserved spermatozoa is an economical, efficient, and safe method for the distribution of mouse strains from one facility to another. However, spermatozoa from some strains, including C57BL/6 (B6), are very sensitive to freezing and thawing and frequently fail to fertilize eggs by conventional in vitro fertilization methods at the recipient mouse facility. Since many genetically engineered mice have the B6 genetic background, this sensitivity poses a major obstacle to studies of mouse genetics. We investigated the feasibility of transporting spermatozoa within epididymides under non-freezing conditions. First, we examined the interval that B6 and B6D2F1 (BDF1) spermatozoa retained their ability to fertilize when stored within epididymides at low temperatures (5 °C or 7 °C). Fertilization rates were >50%, irrespective of the spermatozoa used, when epididymides were stored for 3 d at 7 °C. B6 spermatozoa, but not BDF1 sperm, had better retention of fertilizing ability at 7 °C versus 5 °C. We then transported freshly collected B6 and BDF1 epididymides from a sender colony to a recipient colony using a common package delivery service, during which the temperature was maintained at 5 °C or 7 °C for 2 d. Sufficiently high fertilization rates (68.0–77.5%) were obtained for all experimental groups, except for B6 spermatozoa transported at 5 °C. These spermatozoa were successfully cryopreserved at the recipient facility and, yielded post-thaw fertilization rates of 27.6–66.4%. When embryos derived from the B6 spermatozoa that were transported at 7 °C were transferred into recipient females, 52.7% (38/72) developed to term. In conclusion, transportation of epididymides at refrigerated temperatures is a practical method for the exchange of mouse genetic resources between facilities, especially when these facilities do not specialize in sperm cryopreservation. For the B6 mouse strain, the transportation of epididymides at 7 °C rather than 5 °C, is recommended.